The normal adult alveolar epithelium, comprised of type I (AT1) and type II (AT2) pneumocytes, forms a tight barrier to the passive leak of fluid from the interstitium and vascular compartments into the alveolar space. Maintenance of the relatively dry state of the alveolar spaces depends on the integrity of this barrier, as well as on active Na transport to create an osmotic gradient that drives fluid reabsorption across the alveolar epithelium. Despite the probable major role of AT1 cells (given the large internal surface of the lung that they cover) to alveolar epithelial function, the relative contributions of AT1 vs AT2 cells are not well delineated. The overall goals of this proposal are to evaluate the differential roles of AT1 and AT2 cells in alveolar epithelial function and biology, capitalizing on the recent development of new tools for studying alveolar epithelial (especially AT1) cells. Our central hypotheses are that 1) AT1 cells are important participants in active transepithelial ion transport (and therefore alveolar fluid clearance) across the alveolar epithelium, 2) AT1 cell contributions to alveolar function and biology are reflected in part by their spectrum of gene/protein expression and phenotypic plasticity, and 3) AT1 cells play major roles in the response to alveolar injury and recovery in adult lungs. We will exploit our recent success in obtaining highly purified AT1 cells, our ability to maintain them in primary culture, and availability of AT1 and AT2 cell-specific genes/promoters to conditionally delete ion transporters specifically in either cell type in vivo in order to address these hypotheses by exploring the following Specific Aims: 1) characterize functional properties of rodent AT1 and AT2 cells using isolated AT1 and AT2 cells and monolayers derived from these cells;2) evaluate differential AT1 and AT2 cell contributions to alveolar function by conditional cell-specific knockout of transport genes in transgenic mice in vivo;and, 3) assess AT1 and AT2 cell-specific roles in lung injury and recovery in vitro and in vivo. Results of these studies will provide major new insights into the differential functional and biological roles of AT1 and AT2 cells in normal adult alveolar epithelium and during injury/recovery. Understanding how AT1 and AT2 cells in the adult lung maintain and differentiate their cellular functions, fates and lineages will constitute major advances in alveolar cell biology and could significantly impact the development of novel therapeutic approaches for modulating alveolar homeostasis in health and disease.